Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 9 de 9
Filtrar
Mais filtros










Intervalo de ano de publicação
1.
Vet Parasitol ; 274: 108920, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31493694

RESUMO

Visceral leishmaniasis (VL) is a zoonosis caused by the parasite Leishmania infantum and the dog is its main reservoir in rural and urban areas. The diagnosis of infection is mainly based on the presence of anti-Leishmania IgG antibodies in the serum of infected dogs. In this study, the sensitivity and specificity of qualitative rapid tests (RTs) dual path platform (DPP) Bio-Manguinhos, rapid enzyme-linked immunosorbent assay (ELISA) IDEXX, Kalazar Detect and ALERE, as well as quantitative ELISA Bio-Manguinhos and in-house indirect immunofluorescence assay (IFA) tests were analyzed in sera from infected and uninfected dogs. Serial dilutions of the in-house IFA were compared with RTs and ELISA Bio-Manguinhos. The results showed that none of the tests reached 100% sensitivity and specificity. There was no statistical difference between the analyzed RTs. The most sensitive test was the DPP Bio-Manguinhos (97.9%), while the rapid ELISA IDEXX showed higher specificity (100%). In the treatment setting of infected and/or diseased animals, quantitative tests for monitoring the evolution of antibody titers are required, which indicates the maintenance of in-house IFA in animal handling. Furthermore, we demonstrate that the RTs present higher sensitivity in serum samples with superior antibody titers obtained in the in-house IFA. However, the RTs exhibited false negatives in samples with low titers of antibodies. Among the RTs, only the DPP Bio-Manguinhos presented better performance in this situation. Therefore, the use of RTs for the diagnosis of VL in dogs with low titers of antibodies, such as asymptomatic, should be carefully evaluated.


Assuntos
Doenças do Cão/sangue , Leishmania infantum , Leishmaniose Visceral/veterinária , Testes Sorológicos/veterinária , Animais , Doenças do Cão/diagnóstico , Doenças do Cão/parasitologia , Cães , Leishmaniose Visceral/sangue , Leishmaniose Visceral/diagnóstico , Sensibilidade e Especificidade
2.
PLoS One ; 14(2): e0211831, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30721272

RESUMO

Leishmaniasis encompasses a group of diverse clinical diseases caused by protozoan parasites of the Leishmania genus. This disease is a major public health problem in the New World affecting people exposed in endemic regions. The city of Governador Valadares (Minas Gerais/Brazil) is a re-emerging area for visceral leishmaniasis, with 191 human cases reported from 2008 to 2017 and a lethality rate of 14.7%. The transmission of the parasite occurs intensely in this region with up to 22% of domestic dogs with positive serology for the visceral form. Lu. longipalpis is one of the most abundant sand fly species in this area. Despite this scenario, so far there is no information regarding the circulating Leishmania species in the insect vector Lutzomyia longipalpis in this focus. We collected 616 female Lutzomyia longipalpis sand flies between January and September 2015 in the Vila Parque Ibituruna neighborhood (Governador Valadares/MG), which is located on a transitional area between the sylvatic and urban environments with residences built near a preserved area. After DNA extraction of individual sand flies, the natural Leishmania infections in Lu. longipalpis were detected by conventional PCR, using primers derived from kDNA sequences, specific for L. (Leishmania) or L. (Viannia) subgenus. The sensitivity of these PCR reactions was 0.1 pg of DNA for each Leishmania subgenus and the total infection rate of 16.2% (100 positive specimens). Species-specific PCR detected the presence of multiple Leishmania species in infected Lu. longipalpis specimens in Governador Valadares, including L. amazonensis (n = 3), L. infantum (n = 28), L. (Viannia) spp. (n = 20), coinfections with L. infantum and L. (Viannia) spp. (n = 5), and L. (Leishmania) spp (n = 44). Our results demonstrate that multiple Leishmania species circulate in Lu. longipalpis in Governador Valadares and reveal a potential increasing risk of transmission of the different circulating parasite species. This information reinforces the need for epidemiological and entomological surveillance in this endemic focus, and the development of effective control strategies against leishmaniasis.


Assuntos
Insetos Vetores/parasitologia , Leishmania/classificação , Leishmania/crescimento & desenvolvimento , Psychodidae/parasitologia , Animais , Brasil/epidemiologia , Humanos , Leishmaniose/epidemiologia , Leishmaniose/genética , Leishmaniose/transmissão , Reação em Cadeia da Polimerase , Reforma Urbana
3.
PLoS One ; 14(1): e0211719, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30703138

RESUMO

Visceral leishmaniasis (VL) or kala-azar, the most severe form of leishmaniasis, can lead to death if not properly diagnosed and treated. Correct identification of infected patients and reservoirs is vital for controlling the spread of leishmaniasis. Current diagnostic kits for leishmaniasis show high sensitivity and specificity, but can also result in false negatives and cross reactions with related parasitic infections. New diagnostic methods with greater accuracy are urgently needed for diagnosis of leishmaniasis. In this study, we aimed to uncover a new highly effective antigen for the diagnosis of visceral leishmaniasis in dogs and humans, aiming to improve the accuracy compared with those of current methods of diagnosis. Initially, in-silico epitope prediction analyses identified several potential B-cell epitopes in the repetitive region of Leishmania infantum kinesin, which co-localized with predicted structural disordered regions, suggesting high potential for antigenicity. Based on this analysis, 8.5 genomic motifs, which encode the repetitive sequence of 39 degenerate amino acids, were selected for recombinant expression. BLASTn analysis of this repetitive region indicated that it is absent in the T. cruzi parasite, which is closely related to Leishmania, indicating the specificity of this region. This potentially antigenic protein, named recombinant kinesin degenerated derived repeat (rKDDR), was recombinantly expressed in Escherichia coli BL21-Star using the pET28a-TEV expression vector. We then evaluated the performance of rKDDR in correctly diagnosing Leishmania infection and compared this new assay with currently used diagnostic tests for leishmaniasis. rKDDR showed greater sensitivity and specificity in correctly diagnosing leishmaniasis both in human (sensitivity 92.86% and specificity 100%) and canine (sensitivity 88.54% and specificity 97.30%) sera compared with those of rK39 (human: sensitivity 90.48% and specificity 97.92%; canine: sensitivity 78.13% and specificity 90.09%). In addition, the rKDDR-ELISA outperformed the EIE-LVC kit, which is the serologic kit recommended by the Brazilian Ministry of Health for the diagnosis of canine visceral leishmaniasis. These results indicate that rKDDR is a highly promising candidate for diagnosis of visceral leishmaniasis, and is more accurate than the currently used gold-standard antigens.


Assuntos
Antígenos de Protozoários/sangue , Doenças do Cão/diagnóstico , Cinesina/imunologia , Leishmania donovani/imunologia , Leishmaniose Visceral/diagnóstico , Proteínas Recombinantes/imunologia , Testes Sorológicos/métodos , Sequência de Aminoácidos , Animais , Anticorpos Antiprotozoários/sangue , Anticorpos Antiprotozoários/imunologia , Antígenos de Protozoários/imunologia , Área Sob a Curva , Sequência de Bases , Doenças do Cão/epidemiologia , Doenças do Cão/parasitologia , Cães , Humanos , Leishmania donovani/isolamento & purificação , Leishmaniose Visceral/epidemiologia , Leishmaniose Visceral/parasitologia , Leishmaniose Visceral/veterinária , Estudos Retrospectivos
4.
Front Immunol ; 9: 2535, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30473693

RESUMO

Human ascariasis has a global and cosmopolitan distribution, and has been characterized as the most prevalent neglected tropical disease worldwide. The development of a preventive vaccine is highly desirable to complement current measures required for this parasitic infection control and to reduce chronic childhood morbidities. In the present study, we describe the mechanism of protection elicited by a preventive vaccine against ascariasis. Vaccine efficacy was evaluated after immunization with three different Ascaris suum antigen extracts formulated with monophosphoryl lipid A (MPLA) as an adjuvant: crude extract of adult worm (ExAD); crude extract of adult worm cuticle (CUT); and crude extract of infective larvae (L3) (ExL3). Immunogenicity elicited by immunization was assessed by measuring antibody responses, cytokine production, and influx of tissue inflammatory cells. Vaccine efficacy was evaluated by measuring the reductions in the numbers of larvae in the lungs of immunized BALB/c mice that were challenged with A. suum eggs. Moreover, lung physiology and functionality were tested by spirometry to determine clinical efficacy. Finally, the role of host antibody mediated protection was determined by passive transfer of serum from immunized mice. Significant reductions in the total number of migrating larvae were observed in mice immunized with ExL3 61% (p < 0.001), CUT 59% (p < 0.001), and ExAD 51% (p < 0.01) antigens in comparison with non-immunized mice. For the Ascaris antigen-specific IgG antibody levels, a significant and progressive increase was observed with each round of immunization, in association with a marked increase of IgG1 and IgG3 subclasses. Moreover, a significant increase in concentration of IL-5 and IL-10 (pre-challenge) in the blood and IL-10 in the lung tissue (post-challenge) was induced by CUT immunization. Finally, ExL3 and CUT-immunized mice showed a marked improvement in lung pathology and tissue fibrosis as well as reduced pulmonary dysfunction induced by Ascaris challenge, when compared to non-immunized mice. Moreover, the passive transfer of specific IgG antibodies from ExL3, CUT, and ExAD elicited a protective response in naïve mice, with significant reductions in parasite burdens in lungs of 65, 64, and 64%, respectively. Taken together, these studies indicated that IgG antibodies contribute to protective immunity.


Assuntos
Ascaris suum/imunologia , Imunoglobulina G/imunologia , Substâncias Protetoras/farmacologia , Adjuvantes Imunológicos/farmacologia , Animais , Anticorpos Anti-Helmínticos/imunologia , Antígenos de Helmintos/imunologia , Ascaríase/imunologia , Ascaríase/parasitologia , Feminino , Imunidade/efeitos dos fármacos , Imunidade/imunologia , Imunização/métodos , Interleucina-10/imunologia , Larva/imunologia , Pulmão/imunologia , Pulmão/parasitologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Suínos/imunologia , Suínos/parasitologia , Doenças dos Suínos/imunologia , Doenças dos Suínos/parasitologia , Vacinação/métodos , Vacinas/imunologia
5.
BMC Genomics ; 19(1): 816, 2018 Nov 13.
Artigo em Inglês | MEDLINE | ID: mdl-30424726

RESUMO

BACKGROUND: Trypanosoma cruzi, the etiologic agent of Chagas disease, is currently divided into six discrete typing units (DTUs), named TcI-TcVI. TcII is among the major DTUs enrolled in human infections in South America southern cone, where it is associated with severe cardiac and digestive symptoms. Despite the importance of TcII in Chagas disease epidemiology and pathology, so far, no genome-wide comparisons of the mitochondrial and nuclear genomes of TcII field isolates have been performed to track the variability and evolution of this DTU in endemic regions. RESULTS: In the present work, we have sequenced and compared the whole nuclear and mitochondrial genomes of seven TcII strains isolated from chagasic patients from the central and northeastern regions of Minas Gerais, Brazil, revealing an extensive genetic variability within this DTU. A comparison of the phylogeny based on the nuclear or mitochondrial genomes revealed that the majority of branches were shared by both sequences. The subtle divergences in the branches are probably consequence of mitochondrial introgression events between TcII strains. Two T. cruzi strains isolated from patients living in the central region of Minas Gerais, S15 and S162a, were clustered in the nuclear and mitochondrial phylogeny analysis. These two strains were isolated from the other five by the Espinhaço Mountains, a geographic barrier that could have restricted the traffic of insect vectors during T. cruzi evolution in the Minas Gerais state. Finally, the presence of aneuploidies was evaluated, revealing that all seven TcII strains have a different pattern of chromosomal duplication/loss. CONCLUSIONS: Analysis of genomic variability and aneuploidies suggests that there is significant genomic variability within Minas Gerais TcII strains, which could be exploited by the parasite to allow rapid selection of favorable phenotypes. Also, the aneuploidy patterns vary among T. cruzi strains and does not correlate with the nuclear phylogeny, suggesting that chromosomal duplication/loss are recent and frequent events in the parasite evolution.


Assuntos
Aneuploidia , Doença de Chagas/parasitologia , Variação Genética , Genoma de Protozoário , Proteínas de Protozoários/genética , Trypanosoma cruzi/genética , Sequenciamento Completo do Genoma/métodos , Animais , Doença de Chagas/transmissão , DNA de Protozoário/genética , Genótipo , Humanos , Insetos Vetores/parasitologia , Tipagem Molecular , Filogenia , Trypanosoma cruzi/classificação , Trypanosoma cruzi/isolamento & purificação
6.
Front. Immunol. ; 9: 2535, 2018.
Artigo | Sec. Est. Saúde SP, SESSP-IBPROD, Sec. Est. Saúde SP | ID: but-ib15682

RESUMO

Human ascariasis has a global and cosmopolitan distribution, and has been characterized as the most prevalent neglected tropical disease worldwide. The development of a preventive vaccine is highly desirable to complement current measures required for this parasitic infection control and to reduce chronic childhood morbidities. In the present study, we describe the mechanism of protection elicited by a preventive vaccine against ascariasis. Vaccine efficacy was evaluated after immunization with three different Ascaris suum antigen extracts formulated with monophosphoryl lipid A (MPLA) as an adjuvant: crude extract of adult worm (ExAD); crude extract of adult worm cuticle (CUT); and crude extract of infective larvae (L3) (ExL3). Immunogenicity elicited by immunization was assessed by measuring antibody responses, cytokine production, and influx of tissue inflammatory cells. Vaccine efficacy was evaluated by measuring the reductions in the numbers of larvae in the lungs of immunized BALB/c mice that were challenged with A. suum eggs. Moreover, lung physiology and functionality were tested by spirometry to determine clinical efficacy. Finally, the role of host antibody mediated protection was determined by passive transfer of serum from immunized mice. Significant reductions in the total number of migrating larvae were observed in mice immunized with ExL3 61% (p < 0.001), CUT 59% (p < 0.001), and ExAD 51% (p < 0.01) antigens in comparison with non-immunized mice. For the Ascaris antigen-specific IgG antibody levels, a significant and progressive increase was observed with each round of immunization, in association with a marked increase of IgG1 and IgG3 subclasses. Moreover, a significant increase in concentration of IL-5 and IL-10 (pre-challenge) in the blood and IL-10 in the lung tissue (post-challenge) was induced by CUT immunization. Finally, ExL3 and CUT-immunized mice showed a marked improvement in lung pathology and tissue fibrosis as well as reduced pulmonary dysfunction induced by Ascaris challenge, when compared to non-immunized mice. Moreover, the passive transfer of specific IgG antibodies from ExL3, CUT, and ExAD elicited a protective response in naïve mice, with significant reductions in parasite burdens in lungs of 65, 64, and 64%, respectively. Taken together, these studies indicated that IgG antibodies contribute to protective immunity.

7.
Malar J ; 16(1): 42, 2017 01 24.
Artigo em Inglês | MEDLINE | ID: mdl-28118834

RESUMO

BACKGROUND: The clinical outcome of malaria depends on the delicate balance between pro-inflammatory and immunomodulatory cytokine responses triggered during infection. Despite the numerous reports on characterization of plasma levels of cytokines/chemokines, there is no consensus on the profile of these mediators during blood stage malaria. The identification of acute phase biomarkers might contribute to a better understanding of the disease, allowing the use of more effective therapeutic approaches to prevent the progression towards severe disease. In the present study, the plasma levels of cytokines and chemokines and their association with parasitaemia and number of previous malaria episodes were evaluated in Plasmodium vivax-infected patients during acute and convalescence phase, as well as in healthy donors. METHODS: Samples of plasma were obtained from peripheral blood samples from four different groups: P. vivax-infected, P. vivax-treated, endemic control and malaria-naïve control. The cytokine (IL-6, IL-10, IL-17, IL-27, TGF-ß, IFN-γ and TNF) and chemokine (MCP-1/CCL2, IP-10/CXCL10 and RANTES/CCL5) plasma levels were measured by CBA or ELISA. The network analysis was performed using Spearman correlation coefficient. RESULTS: Plasmodium vivax infection induced a pro-inflammatory response driven by IL-6 and IL-17 associated with an immunomodulatory profile mediated by IL-10 and TGF-ß. In addition, a reduction was observed of IFN-γ plasma levels in P. vivax group. A lower level of IL-27 was observed in endemic control group in comparison to malaria-naïve control group. No significant results were found for IL-12p40 and TNF. It was also observed that P. vivax infection promoted higher levels of MCP-1/CCL2 and IP-10/CXCL10 and lower levels of RANTES/CCL5. The plasma level of IL-10 was elevated in patients with high parasitaemia and with more than five previous malaria episodes. Furthermore, association profile between cytokine and chemokine levels were observed by correlation network analysis indicating signature patterns associated with different parasitaemia levels. CONCLUSIONS: The P. vivax infection triggers a balanced immune response mediated by IL-6 and MCP-1/CCL2, which is modulated by IL-10. In addition, the results indicated that IL-10 plasma levels are influenced by parasitaemia and number of previous malaria episodes.


Assuntos
Citocinas/sangue , Malária Vivax/imunologia , Malária Vivax/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Imunoensaio , Masculino , Pessoa de Meia-Idade , Plasma/química , Adulto Jovem
8.
PLoS Pathog ; 11(12): e1005296, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26641088

RESUMO

Leishmaniasis, a human parasitic disease with manifestations ranging from cutaneous ulcerations to fatal visceral infection, is caused by several Leishmania species. These protozoan parasites replicate as extracellular, flagellated promastigotes in the gut of a sandfly vector and as amastigotes inside the parasitophorous vacuole of vertebrate host macrophages. Amastins are surface glycoproteins encoded by large gene families present in the genomes of several trypanosomatids and highly expressed in the intracellular amastigote stages of Trypanosoma cruzi and Leishmania spp. Here, we showed that the genome of L. braziliensis contains 52 amastin genes belonging to all four previously described amastin subfamilies and that the expression of members of all subfamilies is upregulated in L. braziliensis amastigotes. Although primary sequence alignments showed no homology to any known protein sequence, homology searches based on secondary structure predictions indicate that amastins are related to claudins, a group of proteins that are components of eukaryotic tight junction complexes. By knocking-down the expression of δ-amastins in L. braziliensis, their essential role during infection became evident. δ-amastin knockdown parasites showed impaired growth after in vitro infection of mouse macrophages and completely failed to produce infection when inoculated in BALB/c mice, an attenuated phenotype that was reverted by the re-expression of an RNAi-resistant amastin gene. Further highlighting their essential role in host-parasite interactions, electron microscopy analyses of macrophages infected with amastin knockdown parasites showed significant alterations in the tight contact that is normally observed between the surface of wild type amastigotes and the membrane of the parasitophorous vacuole.


Assuntos
Interações Hospedeiro-Parasita/fisiologia , Leishmania braziliensis/patogenicidade , Leishmaniose Cutânea/genética , Macrófagos/parasitologia , Proteínas de Protozoários/genética , Sequência de Aminoácidos , Animais , Northern Blotting , Western Blotting , Modelos Animais de Doenças , Leishmania braziliensis/genética , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Transfecção , Virulência
9.
BMC Infect Dis ; 15: 35, 2015 Jan 31.
Artigo em Inglês | MEDLINE | ID: mdl-25636730

RESUMO

BACKGROUND: For a long time, the role of CD8(+) T cells in blood-stage malaria was not considered important because erythrocytes do not express major histocompatibility complex (MHC) class I proteins. While recent evidences suggest that CD8(+) T cells may play an important role during the erythrocytic phase of infection by eliminating parasites, CD8(+) T cells might also contribute to modulate the host response through production of regulatory cytokines. Thus, the role of CD8(+) T cells during blood-stage malaria is unclear. Here, we report the phenotypic profiling of CD8(+) T cells subsets from patients with uncomplicated symptomatic P. vivax malaria. METHODS: Blood samples were collected from 20 Plasmodium vivax-infected individuals and 12 healthy individuals. Immunophenotyping was conducted by flow cytometry. Plasma levels of IFN-γ, TNF-α and IL-10 were determined by ELISA/CBA. Unpaired t-test or Mann-Whitney test was used depending on the data distribution. RESULTS: P. vivax-infected subjects had lower percentages and absolute numbers of CD8(+)CD45RA(+) and CD8(+)CD45RO(+) T cells when compared to uninfected individuals (p ≤ 0.0002). A significantly lower absolute number of circulating CD8(+)CD45(+)CCR7(+) cells (p = 0.002) was observed in P. vivax-infected individuals indicating that infection reduces the number of central memory T cells. Cytokine expression was significantly reduced in the naïve T cells from infected individuals compared with negative controls, as shown by lower numbers of IFN-γ(+) (p = 0.001), TNF-α(+) (p < 0.0001) and IL-10(+) (p < 0.0001) CD8(+) T cells. Despite the reduction in the number of CD8(+) memory T cells producing IFN-γ (p < 0.0001), P. vivax-infected individuals demonstrated a significant increase in memory CD8(+)TNF-α(+) (p = 0.016) and CD8(+)IL-10(+) (p = 0.004) cells. Positive correlations were observed between absolute numbers of CD8(+)IL-10(+) and numbers of CD8(+)IFN-γ(+) (p < 0.001) and CD8(+)TNF-α(+) T cells (p ≤ 0.0001). Finally, an increase in the plasma levels of TNF-α (p = 0.017) and IL-10 (p = 0.006) and a decrease in the IFN-γ plasma level (p <0.0001) were observed in the P. vivax-infected individuals. CONCLUSIONS: P. vivax infection reduces the numbers of different subsets of CD8(+) T cells, particularly the memory cells, during blood-stage of infection and enhances the number of CD8(+) memory T cells expressing IL-10, which positively correlates with the number of cells expressing TNF-α and IFN-γ.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Malária Vivax/imunologia , Plasmodium vivax/imunologia , Adulto , Idoso , Contagem de Células Sanguíneas , Estudos de Casos e Controles , Feminino , Citometria de Fluxo , Humanos , Malária Vivax/sangue , Masculino , Pessoa de Meia-Idade , Fenótipo , Adulto Jovem
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA